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Expression and Purification of MC4R/TM2 peptide

등록일
2005년 8월 10일 13시 36분 22초
접수번호
0388
발표코드
24P122포 이곳을 클릭하시면 발표코드에 대한 설명을 보실 수 있습니다.
발표시간
금 <발표Ⅰ>
발표형식
포스터
발표분야
생명화학
저자 및
공동저자
김용애, 박태준, 박종문
한국외국어대학교 화학과,
The melanocortin receptor family consists of five subtypes (MC1R-MC5R) and belongs to the superfamilly of G protein-coupled receptors (GPCRs) which activate the adenylate cyclase signal transduction pathway. Recently, it has been suggested that nomal melanocortin receptor increases energy expenditure and decreases food intake, but genetic disruption of MC4R causes obesity. Therefore, MC4 receptors may be ideal pharmacological targets for treating disorders such as obesity and anorexia. Unfortunately, MC4R is membrane-bound protein that transverse the lipid bilayer of the cell membrane, so it is hard to express and characterize the membrane- bound three-dimensional structure using conventional solution NMR and X-ray crystallography. In this study, we will characterize the membrane-bound three-dimensional structure of MC4R/TM2 peptide using solid-state NMR. We successfully cloned and optimized the expression condition and purified the MC4R/TM2 peptide. PET-31b+ vector and C43(DE3), BL21(DE3) mutant E.coli strain was used. The E.coli cells were grown in M9 minimum media consisting of 15NH4Cl. The fusion protein was purified with a Ni2+-NTA agarose affinity chromatography and chemically cleaved. The yield was 200mg/1L growth with a fusion partner KSI. Finally, we purified MC4R/TM2 peptide using HPLC.

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